Two-sided flow-through immunoassay

ABSTRACT

A two-sided flow-through immunoassay testing device is provided. The device comprises a well having therein a plurality of orifices, the plurality of orifices serving to channel biologic material deposited into the well onto different immunoassay pods, wherein the immunoassay pods may contain immunoassay test layers stacked to create an immunoassay test. The device further includes a results window. Inside the device, and between the window and the pods, there are open sections below each pod to allow a user to view the results of the tests as presented on the reaction layers of the pods through the window.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Application No.62/434,301, filed on Dec. 14, 2016, entitled TWO-SIDED FLOW-THROUGHIMMUNOASSAY (Atty. Dkt. No. RIDL-33396), which is incorporated byreference herein in its entirety.

TECHNICAL FIELD

The following disclosure is related to chromatographic immunoassays and,more specifically, to a two-sided flow-through vertical immunoassaydevice.

BACKGROUND

Chromatographic immunoassays have traditionally used what is known aslateral flow immunoassays. However, lateral flow immunoassays requirethat an analyte migrate across the immunoassay through capillary action,which can take a relatively long time to occur, especially in situationswhere test results are needed as soon as possible. Stacking layers of animmunoassay to allow gravity to pull the analyte through the layersresults in a much quicker and more effective test. Therefore, what isneeded is a vertical two-sided flow-through immunoassay device.

SUMMARY

A two-sided flow-through immunoassay testing device is provided. Thedevice comprises a well having therein a plurality of orifices, theplurality of orifices serving to channel biologic material depositedinto the well onto different immunoassay pods, wherein the immunoassaypods may contain immunoassay test layers stacked to create animmunoassay test. The device further includes a results window. Insidethe device, and between the window and the pods, there are open sectionsbelow each pod to allow a user to view the results of the tests aspresented on the reaction layers of the pods through the window.

BRIEF DESCRIPTION OF THE DRAWINGS

For a more complete understanding, reference is now made to thefollowing description taken in conjunction with the accompanyingDrawings in which:

FIG. 1 illustrates a diagrammatic representation of one embodiment of aimmunoassay test strip;

FIG. 2 illustrates a diagrammatic representation of one embodiment of animmunoassay test wherein an analyte is tested across a plurality of teststrips;

FIG. 3 illustrates a diagrammatic representation of one embodiment of atesting device;

FIG. 4 illustrates a top view of the testing device of FIG. 3;

FIG. 5 illustrates a top view of one embodiment of a testing device;

FIG. 6 illustrates a top view of another embodiment of a testing device;

FIG. 7 illustrates a flowchart of one embodiment of a testing device usemethod;

FIG. 8A illustrates a diagrammatic representation of one embodiment of aprocess for a mobile device application for testing device image captureand image processing, wherein an image alignment indicator is notaligned with the subject of the image;

FIG. 8B illustrates a diagrammatic representation of one embodiment of aprocess for a mobile device application for testing device image captureand image processing, wherein an image alignment indicator is alignedwith the subject of the image;

FIG. 9 illustrates a flowchart of one embodiment of an image analysisprocess using a mobile device;

FIG. 10 illustrates a diagrammatic representation of another embodimentof a process for a mobile device application for testing device imagecapture and image processing, wherein an image alignment indicator isaligned with the subject of the image;

FIG. 11 illustrates one embodiment of a consumer driven biologic anddisease data collection system;

FIG. 12 illustrates one embodiment of a consumer driven biologic anddisease data collection system;

FIG. 13 illustrates an example of a unique biologic ID database table;

FIG. 14 illustrates a flowchart of one embodiment of a biologic datacollection and dissemination process;

FIG. 15 illustrates a perspective view of a system for scanning teststrips;

FIG. 16 illustrates a cross-sectional view of the system of FIG. 15;

FIG. 17 illustrates one embodiment of a vertical flow immunoassaydevice;

FIG. 18 illustrates a cross-sectional view of one embodiment of thevertical immunoassay device of FIG. 17;

FIG. 19 illustrates a color gradient chart;

FIG. 20 illustrates a normalized past tests results rating chart;

FIG. 21 illustrates a mobile device displaying on a screen a mobileapplication variable test functionality;

FIG. 22 illustrates the mobile device of FIG. 21, wherein a housing of atesting device also includes thereon test function indicators;

FIG. 23A illustrates a top view of one embodiment of a two-sidedflow-through immunoassay device;

FIG. 23B illustrates a bottom view of the two-sided flow-throughimmunoassay device of FIG. 23A;

FIG. 23C illustrates a cross-sectional view of the two-sidedflow-through immunoassay device of FIG. 23A-B;

FIG. 24 illustrates one embodiment of an immunoassay pod;

FIG. 25 illustrates another embodiment of an immunoassay pod;

FIG. 26 illustrates one embodiment of a two-sided flow-throughimmunoassay device wherein the device can be opened to gain access tothe inside of the device to install or uninstall pods;

FIG. 27 illustrates a top view of a bottom section of a two-sidedflow-through device;

FIG. 28 illustrates a cross-sectional view of one embodiment of antwo-sided flow-through immunoassay testing device;

FIG. 29A illustrates a top view of another embodiment of a two-sidedflow-through immunoassay device;

FIG. 29B illustrates a cross-sectional view of one embodiment of thetwo-sided flow-through immunoassay device of FIG. 29A;

FIG. 29C illustrates another cross-sectional view of one embodiment ofthe two-sided flow-through immunoassay device of FIGS. 29A-B;

FIG. 29D illustrates another cross-sectional view of one embodiment ofthe two-sided flow-through immunoassay device of FIGS. 29A-C; and

FIG. 29E illustrates a top perspective view of one embodiment of thetwo-sided flow-through immunoassay device of FIGS. 29A-D.

DETAILED DESCRIPTION

Referring now to the drawings, wherein like reference numbers are usedherein to designate like elements throughout, the various views andembodiments of an arbovirus indicative birth defect risk test areillustrated and described, and other possible embodiments are described.The figures are not necessarily drawn to scale, and in some instancesthe drawings have been exaggerated and/or simplified in places forillustrative purposes only. One of ordinary skill in the art willappreciate the many possible applications and variations based on thefollowing examples of possible embodiments.

Referring now to FIG. 1, there is illustrated one embodiment of animmunoassay test strip 100. The test strip 100 is typically housed in atesting device configured to collect a biologic analyte 106 from a userand to direct to the biologic analyte 106 onto the testing strip 100.However, it will be understood that the biologic may be applied onto astrip 100 without the strip 100 needing to be within a testing device.The test strip 100 includes a backing 102. The test strip 100 is made upof multiple sections disposed on the backing 102. A sample pad 104 isdisposed on one end of the strip 100, for collecting the biologicanalyte 106. The biologic analyte 106 may be any biologic needed for usein the immunoassay, such as urine, blood, saliva, stool, sweat, or otherbiologics to be used as an analyte. Various methods may be used toacquire the needed biologic, and such may be provided to the userpackaged with the test, such as swabs, vials, containers, dilutants andother solutions, or any other equipment required. In the case of a bloodanalyte, a few drops of blood may be obtained from a finger stick usinga finger prick device. Such a blood analyte may be blood mixed with anadequate amount of buffered solution to create the sample analyte 106 ora blood sample that is not diluted or otherwise manipulated, in whichcase the blood only is the analyte 106.

The biologic analyte 106, after coming into contact with the sample pad104, begins to migrate across the strip 100 by capillary action, cominginto contact with other sections of the strip 100. A particle conjugatepad 108 is disposed between the sample pad 104 and a test line 110. Theconjugate pad 108 may contain various reagents associated with aparticular antigen, such as a virus, allergen, or bacteria, the reagentsbeing items such antibodies, enzymes, or other reagents needed todiagnose the particular condition. The reagent in the conjugate pad 108may be conjugated with particles of materials such as colloid gold orcolored latex beads. As the analyte 106 migrates through the conjugatepad 108, antibodies present in the sample analyte 106 complex with thereagents in the conjugate pad 108, thereby creating an immune complexthat will migrate to the test zone or test line 110.

The test line 110 (T) may be precoated with the relevant antigen inquestion, i.e., a virus, allergen, or bacteria, for the detection ofantibodies associated with the particular antigen. The immune complexcreated when the analyte 106 passes through the conjugate pad 108 iscaptured onto the antigen contained on the test line 110. This maycreate a qualitative response on the strip where the test line 110 islocated, such as a colored response. In some embodiments, the test line110 may not be a line, but may be other shapes or symbols, such as aplus sign. If no antigen-anti-antigen complexes are present in theanalyte, no reaction occurs in the test line 110 and a qualitativeresponse will not occur.

After passing through the test line 110, the analyte migrates furtheralong the strip to reach a control line 112, where excessanti-antibody-colloidal gold or latex conjugates get bound. Aqualitative response may be shown at the control line 112, indicatingthat the sample has adequately migrated across the testing membrane orsubstrate as intended. It will be understood that the control line 112is not necessarily needed to perform the test, and may be eliminatedentirely, but the control line 112 does provide a comparative examplefor a user reading the test. For example, the control line 112, inembodiments where a colored qualitative response is provided, may appearas an overly saturated color, such as a dark or bright saturated red,once the sample reaches the control line 112. This saturated color maybe used as a comparison against the qualitative response shown on thetest line 110. For example, if the qualitative response shown on thetest line 110 is a much lighter red than that on the test line 110, itmay be that very little reaction occurred at the test line. Of course,if no response is shown at all at the test line 110, no reaction hasoccurred. If the qualitative response at the test line 110 is of asimilar saturation to the control line 112, a strong reaction isindicated.

The strip 100 may not be a continuous substrate. Rather, the varioussections of the strip 100 may be separate from each other, but alladhered to the backing 102. As shown in FIG. 1, the sample pad 104 andthe conjugate pad 108 are separate structures from each other. The testline 100 or zone and the control line 112 or zone are both disposed aspart of a nitrocellulose membrane strip 114. The nitrocellulose membranestrip 114 is also adhered to the backing 102, but separate from thesample pad 104 and the conjugate pad 106. As shown in FIG. 1, the end ofthe sample pad 104 adjacent to the conjugate pad 106 may overlap theconjugate pad 106, with that end of the sample pad 106 lying over theadjacent end of the conjugate pad 106. Similarly, the end of theconjugate pad adjacent to the nitrocellulose membrane strip 114 may lieover the end of the nitrocellulose membrane adjacent to the conjugatepad. This allows for the analyte 106 to be more easily deposited ontoeach section of the strip 100 as it migrates across the strip 100. Afterthe analyte 106 migrates across the nitrocellulose membrane strip 114,and thus across the test line 110 and the control line 112, the analyte106 comes into contact with a wick 116 for absorbtion and collection ofthe analyte 106. The end of the wick 116 adjacent to the nitrocellulosemembrane strip 114 may lie over that adjacent end of the nitrocellulosemembrane strip 114, as shown in FIG. 1.

Several Flow Immune Assays have been directed toward identifyingproteins, molecules of interest, and even immunoglobulins IgG, IgA, andIgM. IgE is an antibody (immunoglobulin E) that is normally present inthe blood freely circulating until it moves into the tissue where it isbound to mast cells through the receptor FcERI (F-C-epsilon-R-one)otherwise known as the high affinity IgE receptor. There is a smallamount of IgE bound to IgE receptors (high and low affinity receptors)on basophils, eosinophils, and other cells in the blood and tissues.

Many assay systems are geared toward the detection of infectiousproteins. All of the aforementioned tests use a non-humanantibody—usually IgG type—e.g., goat IgG antibody directed against aprotein of interest to detect the protein of interest from the sample(blood, urine, saliva, sweat, etc.). This antibody complexes withprotein of interest and forms a complex that travels across the membraneuntil it reaches the test zone. In the test zone there is an IgG typeantibody directed against IgG from that species of animal. As furtherdescribed herein, the present detecting apparatus and method use human(patient/consumer-derived) antibodies from the sample and the test zonethat contains a humanized antibody directed against the protein ofinterest that is preconjugated to a detecting substance that results ina visual change.

Summary of Target Antigen

The target antigens may be proteins, glycoproteins, lipoproteins orother molecular substances capable of eliciting an immune reactionand/or being bound by human specific IgE (sIgE).

Immune Assay to Detect Specific IgE

In the detecting apparatus and method of using the same, the antigensare proteins conjugated to a noble metal, for example, gold, or latexconjugated to antigen in the test zone, for the purpose of detecting thepresence of specific IgE (e.g., anti-peanut IgE in a blood sample from afinger prick). For example, an IgG class antibody (IgG1, IgG2, IgG3, orIgG4) or fragments of those classes of antibodies (fab fragments) whoseorigin may be any animal species (goat, rat, human, etc.) capable ofdetecting human IgE (anti-IgE IgG)—a suitable commercially availablehumanized antibody, such as omaluzimab may be used—may be used to formimmune complexes of IgG-anti-IgE-sIgE that will migrate to the test zonehaving selected specific IgE that can bind to the conjugated antigen.

Immune Assay to Detect Total IgE (not Concerned about Specific IgE)

Another embodiment includes using an IgG class antibody (IgG1, IgG2,IgG3, or IgG4) or fragments of those classes of antibodies (fabfragments) whose origin may be any animal species (goat, rat, human,etc.) capable of detecting human IgE (anti-IgE IgG)—a suitablecommercially available humanized antibody that is preconjugated to adetecting molecule that results in a color change when bound to IgE asthe target antigen in the test zone.

Referring now to FIG. 2, there is illustrated one embodiment of animmunoassay test 200 wherein an analyte 202 is tested across a pluralityof test strips 204. The plurality of test strips 204 may each beconfigured for testing for a particular antigen. For instance, one stripmay be for testing for the presence of streptococcal bacteria (strepthroat), one strip may be for testing for a peanut allergy, one stripmay be for testing for the Zika virus, etc. Additionally, each strip mayalso test for multiple antigens. For example, as shown in FIG. 2,multiple testing panels or lines maybe be incorporated. Each line may befor a particular antigen. As shown in FIG. 2, multiple test lines 206,208, and 208 may be disposed along the plurality of strips 204. A striptesting for allergens may have a panel for testing for peanut allergiesshown at test line 206 (CH1), for cat allergies shown at test line 208(CH2), or grass allergies shown at test line 210 (CH3).

Other examples of configurations for the testing panels can be, but arenot limited to: 1) Food 5: Peanut, milk, soy, wheat, egg; 2) Nut andseed panel: almond, cashew, hazelnut, peanut, pecan, walnut, sesameseed, sunflower seed; 3) seafood: crab, lobster, shrimp, salmon, tuna;4) Pets: cat, dog; 5) Indoor allergens: dust mites, mold mix(alternaria, aspergillus, penicillium, cladosporium), cat, dog; and 6)seasonal allergens: grass (Bermuda, bahia, Johnson, rye, timothy), trees(oak, elm, cedar, mesquite, pine, etc.), weeds (pigweed, ragweed, sage,Russian thistle).

With respect to other non-allergen antigens, the panels may be fortesting for strep, Zika, flu, anthrax, cold viruses, cancer, HPV, Lymedisease, mononucleosis (mono), and other illnesses, and/or otherconditions such as pregnancy (hCG detection) and disease risks. Someembodiments may allow for the testing of various arboviruses(arthropod-borne viruses). Arboviruses are viruses that are transmittedby arthropods, with mosquitos being a common vector for the virus.Vectors are organisms that transfer the virus from a host that carriesthe virus. Thus, in the case of mosquitos, a mosquito that feeds on ahost that is infected with a virus may infect others when that mosquitoagain feeds on an uninfected host. Well-known arboviruses include Denguevirus, Japanese encephalitis virus, Rift Valley fever virus, West Nilevirus, yellow fever virus, chikungunya, and Zika virus. Urine, blood,and saliva and other biologics may be used for arboviruses testing.

Certain antigens or medical conditions may be logically paired together.For instance, a testing device may include both a strip for detection ofpregnancy and a strip for the detection of the zika virus, as the Zikavirus has been known to cause birth defects in infants born to pregnantwomen that are infected with Zika. Thus, combining these two tests intoa single testing device or kit would alert a woman to a potential Zikainfection proximate in time to the time she also discovers she ispregnant, allowing the woman to seek medical attention immediately. Thisis a substantial improvement over past Zika testing, where a woman maybe required to wait weeks before results are returned from a lab afterhaving the biologic collected by her physician. In many cases, this maylead to a woman having passed a state-mandated cutoff point forabortions, such as 24 weeks in some states. Combining a Zika test with apregnancy test and physically linking the two tests, and thus allowingfor a woman to determine a Zika risk at the time of taking a pregnancytest, in which a pregnancy test may be taken as soon as six days afterconception, allows for that woman to take action much sooner than thestate mandated cutoff and waiting for lab results would allow.

Various testing devices that include the test strip 100 or strips may beused, such as a slide that supports the test strip 100, a cassette baseddiagnostic test, a dipstick, or combinations thereof. The test resultsin various embodiments may be in the form of a visual qualitativereading test, a visual semiquantitative format, a reader quantitativeassay format, and/or combinations thereof. Additionally, an electronicimplementation may be used where the result is displayed digitally on ascreen disposed within the apparatus, and visible to the user.

The apparatus and method of detection may be a “one-step” approach fromsample to reading without sample dilution or other sample manipulation.The sample may be diluted or endure other sample manipulation, forexample the blood sample is diluted with a buffer.

Referring now to FIG. 3, there is illustrated a diagrammaticrepresentation of one embodiment of a testing device 300. The testingdevice 300 includes a housing 302 that forms the body of the testingdevice. The housing 302 may be made of plastic, metal, or any materialdurable enough for shipping and subsequent handling by a user. Thehousing 302 may be hollow so that a plurality of test strips 304 may behoused within and so that a biologic may be deposited within the housing302. The testing device 300 may further have a plurality of windows 306,each window being associated with one of the plurality of test strips304, and allowing for a user to view at least the section of thenitrocellulose membrane strip 114 where the test line 110 and controlline 112 are located. The plurality of windows 306 may be open, orcovered with plastic, glass, or other materials that allow for viewingthe plurality of strips 304. A sample well 308 may be disposed on asurface of the housing 302 to allow a user to deposit a biologic intothe housing 302. The sample well 308 would be disposed over or near thesample pad 104 of the test strip or strips 100. In the embodiment shownin FIG. 3, a single sample well 308 is included for collection of asingle type of biologic for testing, with each of the plurality ofstrips 304 being suited for testing for antigens using that particularbiologic sample type. For example, if the testing device 300 is acombined pregnancy and Zika test, having both a pregnancy strip and aZika strip, a urine sample may be deposited into the sample well 308,causing the urine sample to come into contact with the sample pad 104 onboth the pregnancy strip and the Zika strip. It will be understood thatboth of these tests may also be performed with a blood sample.

The testing device 300 may also have disposed on the surface of thehousing a crosshair symbol 310, used as an alignment target. This symbolmay be a graphic printed or adhered to the testing device 300. Thecrosshair symbol 310 is used to align the testing device 300 for thetaking of an image of the testing device 300 using a camera on a mobiledevice, for use in a mobile device application described herein. Inother embodiments, the crosshair symbol 310 may be other types ofsymbols, such as a simple shape (circle, square, etc.), other images(such as a medical cross symbol, an arrow, etc.), or any other type ofimage.

Referring now to FIG. 4, there is illustrated a top view of the testingdevice 300. There is again shown the housing 302, the plurality of teststrips 304, the plurality of windows 306, the sample well 308, and thecrosshair symbol 310.

Referring now to FIG. 5, there is illustrated a top view of oneembodiment of a testing device 500. The testing device 500 includes ahousing 502 having a plurality of test strips 504 within the housing 502and a plurality of windows 506 for display of the plurality of strips504. The housing 502 also includes a plurality of sample wells 508disposed on one side of the testing device 500. Each of the plurality ofsample wells 508 is associated with one of the plurality of test strips504 and each of the plurality of sample wells 508 may be disposed overone of the sample pads 104 on the associated one of the plurality oftest strips 504. This allows for a biologic to be deposited into each ofthe plurality of sample wells 508, with each well 508 serving totransfer the biologic to the test strip underneath the sample well. Thetesting device 500 further includes a crosshair 510. The crosshairsymbol 510 is used to align the testing device 500 for the taking of animage of the testing device 500 using a camera on a mobile device, foruse in a mobile device application described herein.

Referring now to FIG. 6, there is illustrated a top view of anotherembodiment of a testing device 600. The testing device 600 includes ahousing 602 having a plurality of test strips 604 within the housing 602and a plurality of windows 606 for display of the plurality of strips604. The housing 602 also includes a plurality of sample wells 608. Inthis embodiment, the sample wells are located on different ends of thehousing 602. In the case of a two test strip device, the sample wells608 are disposed on opposite ends of the testing device 600. The strips604 would be arranged within the housing in such a way as to allow thesample pad 104 on each of the strip to be disposed underneath one of thesample wells 608. This is useful for testing devices that requiredifferent biological samples. For example, if the testing device 600required a urine sample for one strip and a blood sample for the otherstrip, having the wells 608 disposed on opposite sides of the testingdevice would reduce the likelihood that a urine sample, for instance,might be inadvertently deposited into the well designated for the bloodsample. In embodiments where there are more than two strips, and morethan two wells, the well positions may alternate between the two sidesof the testing device. For instance, a first well for a first stripmight be disposed on the left side of the testing device, a second wellfor a second strip might be disposed on the right side of the testingdevice, a third well for a third strip might be disposed on the leftside of the testing device, a fourth well for a fourth strip might bedisposed on the right side of the testing device, and so on. The testingdevice 600 further includes a crosshair 610. The crosshair symbol 610 isused to align the testing device 600 for the taking of an image of thetesting device 600 using a camera on a mobile device, for use in amobile device application described herein.

The diagnostic test can, for example, be produced in a various formatsfor different users, such as, but not limited to, consumer/in-home usewhere the test is purchased through retail channels which will allowindividuals to get an immediate, cost-effective test result that canlead to specific avoidance and treatment through follow-up with amedical professional.

The diagnostic test can be provided to and used by hospitals and clinicsto provide rapid, on-site test results that are required to prescribecertain medications, such as omaluzimab, by their FDA labels.

This diagnostic assay can be modified to detect the presence of specificIgE in pets.

It is also noted that housing 602 is designed such that both strips 604are disposed in physical proximity thereto and in the same actualhousing. In this manner, both sets are linked physically to each othersuch that they cannot be separated and can be associated with a singleindividual and the actual test cannot be separated. As such, when apatient applies the specimens to the two areas 608 and the test resultsare exhibited, there is a high probability that two tests were performedat the same time associated with the same patient. Additionally, andelectronic chip (not shown) can be embedded within the housing 602 suchthat the housing 602 can be registered to a specific patient andassociated with the medical records of that patient.

Referring now to FIG. 7, there is illustrated a flowchart of oneembodiment of a testing device use method 700. The method 700 begins atstep 702 where a biologic is collected in a sample well or wells of atesting device. The biologic collected may be a non-diluted ornon-manipulated biologic, such as blood, urine, or saliva from the userof the test. Diluted or manipulated biologics may be used instead, asrequired by the specific test. For example, if a viral test requires thebiologic to be added to a solution, the biologic could be added to asolution that has previously been placed in a sterilized vial providedwith the testing device. After the required amount of time has passed,the solution containing the biologic could be deposited into the well orwells. At step 704, the biologic contacts a sample pad disposed on astrip or strips within the testing device. At step 706, the biologicmigrates along the strip or strips to come into contact with a conjugatepad containing antibodies. Antibodies present in the biologic willcomplex with the antibodies in the conjugate pad to create an immunecomplex. At step 708, the biologic migrates into a test zone of thestrip or strips, coming into contact with an antigen. The antibodies inthe conjugate pad serve to provide a means of detection, such as acolored response, if the immune complex binds with the antigen presentin the test zone of the strip. At decision block 710, binding of theantibodies with the antigen may or may not occur depending on ifantibodies associated with the antigen are present in the biologic ornot. If a binding between the antibodies and the antigen does not occurthe process moves to step 712 where no qualitative response is producedon the test line. If a binding does occur, at step 714 a qualitativeresponse is produced on the test line. Whether a binding occurs or not,and whether a qualitative response is produced or not, the process movesto step 716 where the biologic migrates into a control zone of the stripor strips where excess conjugates get bound and produces a qualitativecontrol zone reaction indicating that the sample has adequately migratedacross the testing zone.

It will be understood by one skilled in the art that the antibodies andantigens applied to the testing strip may be altered depending on thetype of condition being tested. For example, in the case of testing formedical conditions that do not involve an illness or infection, likepregnancy, and thus the sample biologic does not contain antibodiesassociated with the condition, antibodies that react to markers beingtested for may be applied to the testing strip instead of an antigen.For instance, pregnancy test requires testing for the presence of hCG.Since hCG is a hormone and not an antibody produced in response to aninfection, the testing strip may have antibodies that will react to thepresence of hCG applied to the testing zone or line of the testingstrip, as well as to the conjugate pad. Similarly, some tests mightrequire antibodies be applied to the testing strip to detect antigenspresent in the sample, rather than antibodies.

Referring now to FIGS. 8A and 8B, there is illustrated a diagrammaticview of one embodiment of a process 800 for a mobile device applicationfor testing device image capture and image processing. The mobile deviceapplication allows for an image of a testing device, such as testingdevice 300, to be captured using a camera installed on a mobile device802 having a screen 804. While the mobile device 802 displays on thescreen 804 the scene captured by the camera, the mobile deviceapplication also displays a graphic on the screen 804 in the form of aboxed outline 806, the size of the outline 806 corresponding to the sizeof the testing device 300. Also displayed on the screen of the mobiledevice 802 within or near the outline is a crosshair graphic 808. A userof the mobile device 802 attempts to align the outline 806 with theborders of the testing device 300 and also attempts to align thecrosshair graphic 808 with the crosshair 310 on the testing device 300.While alignment has not yet been achieved, a misalignment warning 810may appear on the screen of the mobile device 802, indicating to theuser that alignment has not yet been achieved. Such is shown in FIG. 8A.

In FIG. 8B, there is shown the result of a successful alignment of theoutline 806 with the testing device 300 and successful alignment of thecrosshair graphic 808 with the crosshair 310 on the testing device 300.As shown in FIG. 8B, once aligned, a success indicator 812 may appear,such as a check mark or other positive status symbol, on the alignedimage. Successful alignment causes the camera on the mobile device 802to capture the current image of the testing device 300. Other checks mayoccur, including ensuring that the image is focused before the image issaved. This image is then processed to determine a result based on theseverity of the reaction occurring on the test strip. The mobile deviceapplication performs an analysis of the test line captured in the image,counting the number of colored pixels, as well as determining theintensity of the color. The mobile device may then compare this numberand color intensity to that in the control line, providing amathematical evaluation of the test line. Utilizing unique wavelengthsof light for illumination in conjunction with either CMOS or CCDdetection technology, a signal rich image is produced of the test linesto detect the colloid gold or latex particles. This provides anadvantage because a user simply looking at the two lines may not knowwhat the test line indicates, such as when the colored line does appearson the strip, but it is a faded line, rather than a dark line. Based onthis analysis, the mobile device application may provide a resultsindicator 814.

The results indicator 814 may be a qualitative result or a quantitativeresult. For example, and as shown in FIG. 8B, a qualitative result forthe results indicator 814 may indicate, in the case of a testing devicefor testing pregnancy as well as an infection, a plus sign next to aline reading “pregnant:” and a plus sign next to a line reading“infection:” to indicate that the user is both pregnant and infectedwith the bacteria or virus being tested, such as the Zika virus. For aquantitative result, the results might provide a numeric rating. Forinstance, a rating system between 1-100 may be used. If the resultsprovide a low rating to the user, such as a rating of 10, this indicatesa low risk of infection, although medical attention may be sought by theuser anyway. For example, if the user is pregnant, and also receives a10 rating for Zika, this may indicate that Zika was detected in lowamounts. However, the user may still seek medical attention or furthertesting from her doctor because Zika has been known to cause birthdefects. If the rating is a high rating, such as 95, this indicates thatan infection has most likely occurred and medical attention should besought immediately.

This same quantitative rating system may be applied to any test (viralinfections, bacterial infections, pregnancy, and other healthconditions), as the quantitative test can be performed using thesoftware described herein to accurately test bound antibodyconcentrations on the test strip. In some embodiments, a combinedqualitative and quantitative result may be presented, such as both arating and a plus or minus sign being presented, or other types ofquantitative and qualitative indications. Additionally, variouscombinations of tests may be provided for in the testing device, such aspregnancy/Zika, preganancy/flu, pregnancy/strep/Zika, etc.

Referring now to FIG. 9, there is illustrated a flowchart of oneembodiment of an image analysis process 900 using a mobile device. Atstep 902 a mobile device application is launched. At step 904 a cameraon the mobile device is activated and a crosshair indicator and atesting device outline appear on the mobile device screen. At step 906the crosshair indicator presented on the screen of the mobile device isaligned with a crosshair icon on the testing device and the deviceoutline presented on the screen of the mobile device is aligned with theborders of a testing device. At step 908, an indicator of successfulalignment is presented on the screen and an image of the testing deviceis taken by the mobile device camera. At step 910, the mobile deviceapplication processes the image of the testing device to determine testline strength by counting the number of colored pixels contained in thetest line. At step 912, the mobile device application correlates lineintensity with analyte concentrations to further determine test linestrength. At step 914, the mobile device application presents the testresults based on pixel count and line intensity, providing either aqualitative or quantitative result.

In some embodiments, the number of pixels indicating bound antibodies onthe strip may be measured against that in the control line to compareline intensity between the two lines, with the control line acting as anexample of a strong reaction, indicating a strong infection, anddetermining how close the test line intensity is to the control line.This would lead to a logical quantitative result. For instance, if thetest line is determined to have a pixel count and line intensity that is25% of the pixel count and line intensity of the control line, a ratingof 25 may be given. If a qualitative result is to be provided, a ratingof 25 may give a qualitative result that is negative, or it could bepositive depending on the type of condition being tested and knownactual infection results where a rating of 25 occurred for thatcondition.

In some embodiments, the test line may not be compared with the controlline to determine a result. Rather, the mobile device application mayhave access to a database having data on numerous past tests for thesame condition. This data may instead be used as the control. Thisallows the application on the mobile device to retrieve data on pasttests and compare the test line data of the current test to past tests.Overall data for past tests may be provided and compared against, suchas providing an average or a curve of past tests, or individual testsrated as having accurate results may be compared against.

In addition to a status result of an infection or other medicalcondition being provided to the user, other indicators of health mayalso be tested and results thereon provided. This provides for potentialearly identification of pregnancy and risk of morbidity, allowing formedical attention to be sought much more quickly. Indicators of healthmay be detected from biologics, such as urine and blood. Urine, forexample, allows for the detection of glucose levels, proteins, bacteria,and infectious markers. In the case of glucose, glucose is usually notfound in urine, but, if it is, that is an indicator of extremely highlevels of glucose in the body, where the kidneys release excess glucoseinto urine. This is often a sign of diabetes. Protein in the urine mayindicate a malfunctioning of the kidneys, which could be the result ofhigh blood pressure. Similarly, if blood is detected in urine, it couldbe a sign of a problem with the kidneys or the bladder. Blood, forexample, allows for the detection of glucose, inflammation, hormones,genetic defect risks, and metabolic endocrine disorders.

Referring now to FIG. 10, there is illustrated another embodiment of asuccessful alignment of the outline 806 with the testing device 300 andsuccessful alignment of the crosshair graphic 808 with the crosshair 310on the testing device 300, wherein quantitative results for health riskindicators are provided. In this embodiment, the results indicator 814provides a qualitative result for pregnancy, and quantitative resultsfor other health risk indicators. In the embodiment shown in FIG. 10,the health risk indicators being tested are markers for blood pressureand for glucose levels. For blood pressure, this is a test for markersin the blood that can be associated with high blood pressure. Thesecould be test for such things as low levels of vitamin D and the such.Studies have shown that patients suffering from essential hypertensionwill be under oxidative stress and Malondialdehyde (MDA) is theprincipal and most studied product of polyunsaturated fatty acidpre-oxidation. This can show an indirect correlation with anti-oxidants,particularly with superoxide dismutases (SODs) (r=0.573) and catalase(r=0.633) representative anti-oxidant are involved in reducing thestress of a patient's biological system during hypertension. Anothermarker for hypertension is buildup of uric acid, where in uric acid is amarker for xanthine oxidase-associated oxidants and that the lattercould be driving the hypertensive response. Additional markers arecortisol, a hormone. The test strips 604 can test for the differentbiological markers.

The results indicator 814 provides numeric ratings, in this case, 1-100,with the blood pressure rating being 88 and the glucose rating being 95.This indicates that both blood pressure and glucose are extremely high.Due to this, an additional alert indicator 1002 is presented to the useron the screen of the mobile device, alerting the user to seek medicalattention immediately. This is to ensure that the health of both thepregnant woman and the fetus can be checked as close to the time ofpregnancy detection as possible and medical attention received ifneeded.

Referring now to FIG. 11, there is provided a flowchart of oneembodiment of a pregnancy disease risk assessment process 1100. Theprocess 1100 begins at step 1102 where a biologic is collected anddeposited in a testing device for testing of the biologic. At step 1104,the biologic is processed by the testing device for detection ofpregnancy and various other medical conditions. These medical conditionsmay be high blood pressure, diabetes, bacterial or viral infections,inflammation, an increase in hormone levels, genetic disease markers,and/or metabolic disorders. At step 1106, a mobile device is used tocapture an image of the testing device after testing is complete. Insome embodiments, test results may be immediate. In other embodiments,and depending on the medical conditions being tested, the test may takea certain amount of time to complete. In this case, the user of the testwould be alerted to this fact in instructions provided with the testingdevice. Additionally, a visual indicator on the testing device may alertthe user that a test is now complete. At step 1108, the mobile deviceprovides a rating for each medical condition being tested, such as thatdescribed with respect to FIG. 10 herein.

At decision block 1110, it is determined whether the ratings for eachcondition exceed a certain threshold for that condition. If not, theprocess 1100 moves to step 1112, where an indication is presented to theuser via the mobile device screen that medical attention is notcurrently advised or necessary. If at step 1110 it is determined that atleast one of the medical conditions being tested rises above a certainthreshold, the process 1100 moves to step 1114 where a warning ispresented to the user via the mobile device screen that medicalattention is advised. The thresholds for medical conditions may nottrigger a warning even if a rating exceeds a threshold, if, in the eventof multiple tests being performed, the combined test results do notwarrant immediate medical attention. For example, if a user is testingfor a cold virus, blood pressure, and glucose, and only the cold virusrating is above the threshold, there may not be a warning provided tothe user. Additionally, ratings may be weighted or aggregated based onthe medical conditions being tested. For example, if blood pressure,inflammation, and glucose are being tested for, and they all are givenonly moderate ratings that do not rise above the threshold for anycondition individually, an warning to seek medical attention may stillbe provided due to the combination of conditions taken together.

Referring now to FIG. 12, there is illustrated one embodiment of aconsumer driven biologic and disease data collection system 1200. Datacollected from users performing the tests disclosed herein effectivelycan provide a wealth of information. As tests are performed data may bepassed by a plurality of mobile devices 1202 being used by usersperforming tests to a database 1204, the database being at a remoteserver 1206, over a network 1208. The user is sourcing a biologic fromuser's own body. This is done at home, not in a lab, hospital, orclinic. Each particular test would expect a certain type of biologic tobe provided. For instance, for a pregnancy test, a urine sample isprovided and tested for pregnancy markers. For a stool test, the stoolmight be dissolved in a vial with a solution provided with the testingdevice/kit, and tested for various disease or infectious markers. Dataand results from the tests may be stored on the database 1204 at theremote server 1206. As described herein, this data may be used as acontrol for testing analysis for users of the plurality of mobiledevices 1202. This data may also be used to provide data sets forbiologics to a medical organization 1210. The medical organization 1210may be doctor's offices, researchers, hospitals, testing labs, and otherindividuals or organizations that have an interest in the health andanalysis of users taking the test and of their biologic samples. In thisway, data can be gathered from a variety of biologics tested for avariety of different medical conditions and characteristics.

Referring now to FIG. 13, there is illustrated an example of a uniquebiologic ID database table 1300. The table 1300 is illustrative of thetype of data stored in association with data for a biologic transmittedby the plurality of mobile devices 1202 for storage on the database1204. A biologic ID header 1302 is provided that shows that the biologicsample has been given a unique ID. All data concerning the biologic maybe stored in association with the unique biologic ID. The table 1300also includes a biologic type entry 1304. This designates what type ofbiologic that the biologic associated with the unique ID is, such asblood, urine, stool, saliva, sweat, or other biologics. The table 1300also provides a plurality of test ratings 1304, for various testsperformed on the biologic. In the example shown in FIG. 13, a bloodbiologic is provided having an assigned ID of 2402, and having beentesting for pregnancy markers, the Zika virus, and for glucose levels.The rating for pregnancy was a 99 rating, the rating for a Zikainfection was a 75, and the rating for glucose levels was a 10. Thiswould indicate that the test subject has an extremely high likelihood ofboth a pregnancy and a Zika infection, which would have resulted in awarning to seek medical attention at the conclusion of the tests. Otherinformation may also be stored in the database in relation to thebiologic, including other condition ratings, time and date each test wasperformed, user information such as ethnicity, gender, and age, andstatus indicators such as whether a test subject visited a physician asa result of the tests. The database 1204 thus provides the test subjectwith a growing collection of information that may be accessed by thetest subject. This allows the test subject to present the test resultsto her physician for medical attention or additional testing, and allowsfor others who may access the database, such as disease researchers, tohave access to data on various biologic samples and their markers.

Referring now to FIG. 14, there is illustrated a flowchart of oneembodiment of a biologic data collection and dissemination process 1400.The process 1400 begins at step 1402 where a user of a testing devicecollects a biologic sample for use in a test or a series of tests. Atstep 1404, the test or series of tests are performed on the biologicsample. At step 1406, a mobile device application checks the biologicsample the testing device result to determine a quantitative result ofthe test to provide a correlative value for the condition being testedin the biologic sample. At step 1408, the test results and correlativevalues, or multiple values if multiple tests on the biologic sample wereconducted, are transmitted to the remote server 1206. At step 1410, thebiologic sample is given a unique identification number in the database1204. At step 1412, the test results and correlative value or values arestored in the database 1204 in association with the uniqueidentification number given to the biologic sample collected and inassociation with the particular tests performed. This way, theparticular biologic sample may have various characteristics stored andretrieved in association with the biologic sample, and the test resultsmay also be retrieved when data on a particular test is needed on across-section of users.

At step 1414, the results are provided to the user on the user's mobiledevice. At step 1416, the results are provided to the user's healthcareprovider. The healthcare provider may receive the test results due to avisit from the user, the user bringing the results of the test with heron her mobile device, or the healthcare provider may receive the resultsfrom the database 1204 if the healthcare provider has permission toaccess the database 1204, or if access is granted in anticipation of theuser's appointment with the healthcare provider. At step 1418, the testresults are also provided to other healthcare industry individuals andorganizations, including medical researchers, hospitals, and others.

Referring now to FIG. 15, there is illustrated a perspective view of asystem for scanning test strips. The housing 604, as describedhereinabove with respect to FIG. 6, is illustrated as being disposedwithin a slot 1502 in a test housing 1504. The test housing 1504 isconnected through a line 1506 to a PC 1508. When the housing 604containing the test strips 604 after being subjected to the biologics isinserted within the slot 1502, the test housing 1504 will scan the teststrips 604 and analyze the results with the PC 1508. The PC 1508 willrun some type of algorithm that can analyze the results of both of thetest strips 604. There can be provided to indicators 1510 and 1512 onthe surface of the test housing 1504, one being, for example, a readyLED and one being a green LED. The PC 1508, after analyzing results, canthen provide a warning indicator such as lighting up the green LED for apositive indication of pregnancy and the red LED for indicating thatthere is some issue. As an example, suppose that the second test striptested for the Zika virus. If so, a warning would be appropriate tooutput and activate the red LED. There could be any other type ofindicator associated with the second test at 604 that, in a combinationwith the test results of the first test strip, i.e. for testing for thepresence of a pregnancy state, testing for such things as diabetes, etc.Further, although only two test strips 604 are illustrated, there couldbe multiple test strips testing for many different biological issuesthat may be present in an individual. In this embodiment, by insertingthe housing 602 into the test housing 1504 and allowing the PC 1508 toanalyze the results, the indicators associated with the test strips canbe analyzed with the assumption that all of the test return results wereassociated with an individual and in proximate time to each other. Thatmeans that the individual patient applied biological specimens, such asurine, blood, etc., to the appropriate test strips and, since these wereall contained within the same test strip housing 602, this provides anindication that they are associated with a single patient. Further, thetest performed will typically be a test that will provide a veryshort-term response. Thus, the specimens can be applied to the teststrips 604 in the test strip housing 602 and then inserted within theslot 1502 for testing by the PC 1508.

Referring now to FIG. 16, there is illustrated a cross-section of thetest housing 1504. It can be seen that the test strip housing 1602 ispassed in slot 1502 past the camera 1602. The camera 1602 is operable toscan a small cross-section of the test strips 604 on the surface thereofas the test strip housing 602 passes thereby. Of course, there couldalso be a much larger camera provided for taking an entire image of thetest strips 604 after being inserted within the test housing 1504. Thecamera 1602 is connected via a wire 1604 two in electronics package 1606to process the information and send it to the PC 1508. The electronicspackage 1606 will also drive the indicators 1510 and 1512.

Referring now to FIG. 17, there is illustrated one embodiment of avertical flow immunoassay device 1700. It will be understood thattesting device 300 and other embodiments herein illustrate a lateralflow immunoassay device. However, other types of immunoassay devices maybe used. For example, vertical flow immunoassay devices may be used, atwo-sided flow through assay, or even a sandwich ELISA test may be run.

The testing device 1700 includes a housing 1702 that forms the body ofthe testing device. The housing 1702 may be made of plastic, metal, orany material durable enough for shipping and subsequent handling by auser. The housing 1702 may be hollow so that a plurality of immunoassaytest pads 1704 may be housed within and so that a biologic may bedeposited within the housing 1702. The testing device 1700 may furtherhave a plurality of sample wells 1706, each sample well having one ofthe plurality of immunoassay test pads 1704 disposed within, andallowing for a user to view at least a section of a nitrocellulosemembrane of each of the immunoassay test pads 1704, the membrane 1708having a test line 1708 and control line 1710. The testing device 1700may also have disposed on the surface of the housing a crosshair symbol1712, used as an alignment target. This symbol may be a graphic printedor adhered to the testing device 1700. The crosshair symbol 1712 is usedto align the testing device 1700 for the taking of an image of thetesting device 1700 using a camera on a mobile device, for use in amobile device application described herein. In other embodiments, thecrosshair symbol 1712 may be other types of symbols, such as a simpleshape (circle, square, etc.), other images (such as a medical crosssymbol, an arrow, etc.), or any other type of image. In otherembodiments, the device 1700 may be configured in such a way as to allowa biologic sample to be deposited into a sample well, and to present theresults of the test on the opposite side of the housing. Such aconfiguration would allow the biologic to flow through the testing padwithin the housing, with the reaction occurring on a reactive membraneon the side of the device opposite the sample well, with the devicehaving a window for viewing the results.

Referring now to FIG. 18, there is illustrated a cross-sectional view ofone embodiment of the vertical immunoassay device 1700. There is shownone of the plurality of immunoassay test pads 1704 residing within thehousing 1702 below one of the plurality of sample wells 1706. The one ofthe plurality of immunoassay test pads 1704 includes a immunoreactivemembrane 1802, such as the nitrocellulose membranes disclosed herein.The immunoreactive membrane 1802 may have particle conjugates disposedthereon that binds when a biologic sample is received onto theimmunoreactive membrane 1802 via the sample well 1706, if the biologicsample contains the antigens or antibodies, or other indicators, forwhich the test is configured. The one of the plurality of immunoassaytest pads 1704 also includes an absorbent pad 1804 for collection ofexcess biologic sample. It will be understood that the cross-sectionalview illustrated in FIG. 18 shows one well of the plurality of samplewells 1704. The other wells included in the device 1700 would beconfigured in a similar manner as that shown in FIG. 18. There may alsobe included in the device 1700 an inner separating wall between each ofthe plurality of immunoassay test pads 1704, to ensure that excessbiologic material that is not adequately absorbed by the absorbent pad1804 does not contaminate neighboring immunoassay test pads.

Referring now to FIG. 19, there is illustrated a color gradient chart1900. When the mobile application described herein captures an image ofthe testing device, in some embodiments each pixel that makes up thetest line captured in the image is processed to place it on a colorgradient scale. In some embodiments, this placement may be done byexamining the RGB values of the pixel. For any given test, there may bea visual color indicator (such as a test line) presented to the user ofthe test to indicate whether a reaction occurred. The color that is tobe presented is known for the given test. Additionally, in someembodiments, the strength of the reaction will affect the strength ofthe color indicator. For example, if a test is supposed to produce abrown colored indicator, an image can be taken of the colored indicatorto examine each pixel of the colored indicator to determine the strengthof the color produced on the testing device, which indicates thestrength of the reaction, and thus the risk level for the user.

The embodiment illustrated in FIG. 19 uses as an example a set of pixelRGB results for a test that produces a red colored indicator on the teststrip when a reaction has occurred. There can be seen an origin point1902 on the chart 1900, wherein the RGB value is (255, 255, 255) orwhite. This may represent a no reaction state for the test strip, sincethe test line on the strip may only appear as a white blank space if noreaction has occurred. An x axis 1904 represents the color green,wherein the amount of green in the pixel decreases as it moves away fromthe origin in relation to the x axis 1904. A y axis 1906 represents thecolor blue, wherein the amount of blue in the pixel decreases as itmoves away from the origin in relation to the y axis 1906. A diagonalline 1908 running in between the x axis 1904 and the y axis 1906represents the color red, wherein the diagonal line 1908 running throughthe center of the chart 1900 is a maximum red color all along thediagonal line 1908. If a pixel has less red than a 255 value, the pixelwould be plotted away from the diagonal line 1908 in relation towhichever color is more predominant. For instance, if the pixel has RGBvalues of (127, 50, 205), a shade of purple, the pixel would be plottedsomewhere in the lower right quadrant of the chart 1900. FIG. 19 furtherillustrates an example plurality of pixel plot points 1910, connected bya curved line, wherein the example plurality of pixel plot points 1910shows tests results that likely indicate a positive reaction, as theplot points are all located near the diagonal line 1908, demonstratingthat the colored indicator was a heavy red color for the most part.

Referring now to FIG. 20, there is illustrated a normalized past testsresults chart 2000. The captured pixels may be normalized into a singlevalue for determining whether there is a likelihood of infection,pregnancy, or whatever else the test is designed to detect. This may bedone in various ways. For example, the shade of red in all the pixelsmay be averaged to reach a single RGB value. Outliers may be left out sothat the average is not heavily skewed, especially when there are fewoutliers present. This RGB value may then be given a value, such as arisk rating, ranging from 0 to 100. For example, an RGB value of (255,255, 255) would be given a rating of 0. An RGB value of (255, 0, 0)would be given a rating of 100. An RGB value of (205, 150, 75) may begiven a rating of 70, and so on. This normalized value may then be usedto compare the user of the test to users of past tests to determine arisk level. In some embodiments, the control line and the test line maybe captured and the results compared, as well. In addition, the realresults of risk levels may also be used to adjust the stored normalizedvalue. For instance, if a particular RGB value that seems to indicate astrong reaction repeatedly was found to not indicate an infection, thisvalue may be adjusted to provide a lower risk rating. For instance, if aphysician who saw a patient who had a (205, 150, 75) RGB value laterreported to the operator of the server 1206 that further testing showedno infection was present, and if this trend continued substantially asreported by other physicians or medical organizations, subsequent testresults by other test users that were near the RGB value of (205, 150,75) may be given a lower rating.

Chart 2000 illustrates how past tests results may be collected and usedto determine the risk of a current test user. A y axis 2002 represents arisk level rating, ranging from 0 at the origin to 100. An x axis 2004represents time, wherein a plurality of normalized test results isplotted on the chart 2000. The chart 2000 is further divided intosections across the y axis 2002, indicating various risk levelthresholds. For instance, and as illustrated in the chart 2000, theremay be at certain rating levels different thresholds of risk labeled aslow, moderate, above average, and high risk levels. These thresholds maybe moved over time as more data is accumulated via users conductingtests and the mobile application storing the data on the tests. When auser conducts a test, the user's normalized rating can be plottedsimilarly to past test results and weighed against them in order toprovide a risk level for the user.

Referring now to FIG. 21, there is illustrated the mobile device 802displaying on the screen 804 a mobile application variable testfunctionality. There is displayed on the screen 804 a plurality of testfunctions 2102. The plurality of test functions 2102 may be buttons thatcan be selected by a user to switch between tests within the mobileapplication. This allows for all test functions to be within the samemobile application. For each test run by the mobile application, datafor the particular test chosen is utilized in performing the test,pulling the data from the remote server 1206.

Referring now to FIG. 22, there is illustrated the mobile device 802 ofFIG. 8B, wherein the housing 302 of the testing device 300 also includesthereon test function indicators 2202 and 2204. The test functionindicators 2202 and 2204 are visual markers located on the housing 302that identify to the mobile application the types of tests for which thetesting device 300 is configured. These indicators may be any symbol,alphanumeric character, shape, etc. that can be added to the surface ofthe testing device 300. The mobile application is programmed torecognize the indicator and perform the test function associated withthe indicator. For example, the embodiment illustrated in FIG. 22 showsa “P” symbol for test function indicator 2202 and a “Z” symbol for testfunction indicator 2204. In this embodiment, test function indicator2202 indicates that one test strip in the testing device 300 is apregnancy test, while test function indicator 2204 indicates that onetest strip in the testing device 300 is a Zika test. This is used formerely illustrative purposes, and any recognizable symbol may be usedfor these two test functions, and any other type of test may have asymbol assigned in this way as well. Further, in some embodiments theremay only be one indicator on the housing 302, even if there are multipletests. This single indicator would be for the combined test. Forexample, if the testing device 300 of FIG. 22 had a single symbol of“PZ,” this would indicate that the testing device 300 is a combinedpregnancy and Zika testing device, allowing for the mobile applicationto recognize such and perform each test with knowledge of which strip isassociated with which test based on the stored data on the testingdevice associated with the “PZ” symbol.

Referring now to FIG. 23A, there is illustrated a top view of oneembodiment of a two-sided flow-through immunoassay device 2300. The topof the device 2300 includes a sample well 2302 having a plurality oforifices 2304. The sample well 2302 has a solid bottom, with the solidbottom have the plurality of orifices 2304 disposed therein. Theplurality of orifices 2304 allow for a biologic sample to be introducedinto the sample well 2302, with the biologic sample entering each one ofthe plurality of orifices 2304 to divert the biologic sample downdifferent paths of the device 2300. These different paths allow formultiple tests to be run on the same biologic sample. Each one of theplurality of orifices 2304 may lead to an enclosed area within thedevice 2300 that contains a specific set immunoassay layers. Eachenclosed area below each of the plurality of orifices 2304 may becompletely closed off from the other enclosed areas so that the biologicsample does not enter into any enclosed area from another enclosed area,contaminating the test being performed within the enclosed area. Thedevice 2300 may also have on the top of the device 2300 a batterycompartment 2306. The battery compartment 2306 allows for a battery tobe inserted into the battery compartment 2306, to supply a positive ornegative charge to the biologic sample deposited within the device 2300.

Thus, the device also allows for immunoelectrophoresis to be performed.Immunoelectrophoresis is a powerful analytical technique with highresolving power as it combines separation of antigens by electrophoresiswith immunodiffusion against an antiserum. The increased resolution isof benefit in the immunological examination of serum proteins.Immunoelectrophoresis aids in the diagnosis and evaluation of thetherapeutic response in many disease states affecting the immune system.It is usually requested when a different type of electrophoresis, calleda serum protein electrophoresis, has indicated a rise at theimmunoglobulin level. Immunoelectrophoresis is also used frequently todiagnose multiple myeloma, a disease affecting the bone marrow. Agaroseas 1% gel slabs of about 1 mm thickness buffered at high pH (around 8.6)is traditionally preferred for the electrophoresis as well as thereaction with antibodies. The agarose was chosen as the gel matrixbecause it has large pores allowing free passage and separation ofproteins, but provides an anchor for the immunoprecipitates of proteinand specific antibodies. The high pH was chosen because antibodies arepractically immobile at high pH. An electrophoresis equipment with ahorizontal cooling plate was normally recommended for theelectrophoresis.

Immunoprecipitates may be seen in the wet agarose gel, but are stainedwith protein stains like Coomassie Brilliant Blue in the dried gel. Incontrast to SDS-gel electrophoresis, the electrophoresis in agaroseallows native conditions, preserving the native structure and activitiesof the proteins under investigation, therefore immunoelectrophoresisallows characterization of enzyme activities and ligand binding etc. inaddition to electrophoretic separation. The immunoelectrophoreticanalysis ad modum Grabar is the classical method ofimmunoelectrophoresis. Proteins are separated by electrophoresis, thenantibodies are applied in a trough next to the separated proteins andimmunoprecipitates are formed after a period of diffusion of theseparated proteins and antibodies against each other. The introductionof the immunoelectrophoretic analysis gave a great boost to proteinchemistry, some of the very first results were the resolution ofproteins in biological fluids and biological extracts. Among theimportant observations made were the great number of different proteinsin serum, the existence of several immunoglobulin classes and theirelectrophoretic heterogeneity.

Two factors determine that immunoelectrophoretic methods are not widelyused. First they are rather work intensive and require some manualexpertise. Second they require rather large amounts of polyclonalantibodies. Today gel electrophoresis followed by electroblotting is thepreferred method for protein characterization because of its ease ofoperation, its high sensitivity, and its low requirement for specificantibodies. In addition proteins are separated by gel electrophoresis onthe basis of their apparent molecular weight, which is not accomplishedby immunoelectrophoresis, but nevertheless immunoelectrophoretic methodsare still useful when non-reducing conditions are needed. To accomplishimmunoelectrophoresis in the device 2300, certain layers of theimmunoassay may be a gel, such as Agarose, into which the antibodies ofthe biologic sample may be introduced.

Referring now to FIG. 23B, there is illustrated a bottom view of oneembodiment of the two-sided flow-through immunoassay device 2300. Thebottom of the device 2300 includes a test results window 2308. The testresults window 2308 allows for a plurality of results indicators 2310 tobe viewable to a user of the device 2300. As a biologic sample soaksthrough each layer of the immunoassay, the user can then see through thesoaked layers, including a transparent wicking layer, to see the resultson a reaction membrane. The plurality of test results indicators 2310may be test lines, such as those described with respect to otherimmunoassays herein, or may be simple dot indicators, which functionsimilarly to the test lines. As can be seen in the example shown in FIG.23B, two out of three test results indicators are displaying results,these two being oriented on the left side of the test results window2308 and designated with a ‘T’ near the area of the test results window2308 where the two indicators appear. A third area designated with a ‘T’in the upper right portion of the test results window 2308 is shown tonot display a result. This is to demonstrate that test results may notappear if the test performed on the biologic does not produce a reactionand therefore a positive result.

The test results window 2308 also includes a control indicator 2312. Thecontrol indicator 2312 indicates that the biologic has completely flowedthrough the device 2300 and is used to compare the plurality of testresults indicators 2310 against the control indicator 2312. The bottomof the device 2300 also includes a vacuum port 2314. The vacuum port2314 allows for a suction device to be attached to the device 2300 topull the biologic sample through the device 2300 at a faster rate thangravity would allow, and to pull more biologic sample through than wouldotherwise move through the device 2300. The bottom of the device 2300also includes a crosshair indicator 2316, for use with a mobile deviceand the application and system described herein to align the device 2300with a camera to perform testing analysis on the results provided by thedevice 2300.

Referring now to FIG. 23C, there is illustrated a cross-sectional viewof one embodiment of the two-sided flow-through immunoassay device 2300.The device 2300 further includes a plurality of immunoassay stackssituated underneath each of the plurality of orifices 2304. Each of theplurality of immunoassay stacks are enclosed separately from each otherimmunoassay stacks. Each of the plurality of immunoassay stacks are fortesting a specific condition, infection, etc. The device 2300 allows forthe insertion of different immunoassay stacks, or pods, into the body,or cassette, of the device 2300. This allow for a custom array of testsin the form of pods to be inserted into the cassette. In someembodiments, a single pod would be the entire immunoassay stack thatincludes all the layers needed to perform a particular test.

For example, as shown in FIG. 23C the pods each have multiple layers,but the entire pod is inserted into the cassette. FIG. 23C shows examplelayers that may be included in the pods. Such layers may include aliquid permeable membrane layer 2318. This layer will only allow liquidmaterial to enter further into each of the plurality of orifices 2304.The pods may also include an opaque layer 2320 which allows for thebiologic sample to pass through, but prevents the user of the devicefrom seeing, either through the sample well 2302 or the test resultswindow 2308, all the way through the device from either end. The podswill also include a reaction membrane 2322, which is the layer whereconjugates may be bound if the proper reaction takes place. The pods mayalso include wicking layers 2324 located at the bottom of each pod forabsorption of excess biologic material. In some embodiments, each podmay have its own wicking layer within the pod, as shown in FIG. 23C. Inother embodiments, the wicking layer may be a separate layer outside ofthe pod, which collects biologic material from all of the pods. It willbe understood by one skilled in the art that other layers may beincluded in a pod as required by the particular tests being performed.For example, there is also shown in FIG. 23C a blocking layer 2326.Blocking layers have a blocking agent applied thereto to filter outunneeded material from the biologic sample to produce more focused testresults. Such a blocking agent often used is milk, or bovine serumalbumin, but any agent needed for the test may be used.

There is additionally shown in FIG. 23C a plurality of electrodes 2330extending from the first battery compartment 2306 and the second batterycompartment 2316, to each of the pods, in order to provide a positive ornegative current to the pods if needed for the particular tests beingperformed.

Referring now to FIG. 24, there is illustrated one embodiment of animmunoassay pod 2400. The pod 2400 includes an immunoassay layer 2402within a housing 2404. The housing 2404 surrounds the immunoassay layer2402, while still allowing for biologic material to enter the housing2404 so that it comes into contact with the immunoassay layer 2402, andallowing for biologic material to leave the housing 2404 so that it cancontact layers below the pod. To accomplish such, the housing 2404 hason the bottom side a first lip 2406 and on the top side a second lip2408. The lips 2406 and 2408 hold the immunoassay layer 2402 in place sothat it does not fall out of the open areas on the top and bottom sidesof the housing 2404, while still allowing biologic material to passthrough the open areas. The housing 2404 may also have a front end 2410and a back end 2412. The front end 2410 and the back end 2412 may befully enclosed ends such that the only open areas are on the top andbottom sides, so that the immunoassay layer 2402 is more secure and sothat biologic material does not leak out of the front end 2410 or theback end 2412. It will be understood that the pod of FIG. 24 may includeone layer, more than one layer, or all the layers needed for aparticular immunoassay test. To include multiple layers, each layerwould be stacked within a single housing 2404. The housing 2404 may bemade of any appropriate material, such as plastic.

Referring now to FIG. 25, there is illustrated another embodiment of animmunoassay pod 2500. The pod 2500 includes a plurality of immunoassaylayers 2502 within a housing 2504. The housing 2504 surrounds theplurality of immunoassay layers 2502, while still allowing for biologicmaterial to enter the housing 2504 so that it comes into contact withthe plurality of immunoassay layers 2502, and allowing for biologicmaterial to leave the housing 2504 so that it can contact layers belowthe pod. To accomplish such, the housing 2504 has on the bottom side afirst lip 2506 and on the top side a second lip 2508. The lips 2506 and2508 hold the plurality of immunoassay layers 2502 in place so that itdoes not fall out of the open areas on the top and bottom sides of thehousing 2504, while still allowing biologic material to pass through theopen areas. The housing 2504 may also have a front end 2510 and a backend 2512. The front end 2510 and the back end 2512 may be fully enclosedends such that the only open areas are on the top and bottom sides, sothat the plurality of immunoassay layers 2502 is more secure and so thatbiologic material does not leak out of the front end 2510 or the backend 2512. It will be understood that the pod of FIG. 25 any number oflayers as needed for a particular immunoassay test. The housing 2504 maybe made of any appropriate material, such as plastic.

Referring now to FIG. 26, there is illustrated one embodiment of atwo-sided flow-through immunoassay device 2600 wherein the device 2600can be opened to gain access to the inside of the device 2600 to installor uninstall pods. There is shown a top section 2602 having a prong 2604of each end thereof. The prongs 2604 fit into a bottom section 2606 tolatch and secure the device 2600 together.

Referring now to FIG. 27, there is illustrated a top view of a bottomsection 2702 of a two-sided flow-through device 2700. The view shown inFIG. 27 is of the bottom section 2702, such as that described withrespect to FIG. 26, with the top section removed and viewing the insideof the device 2700. There exists a cavity 2704 having a housing 2706 forinstalling a plurality of immunoassay pods 2708. The pods are insertedinto the cavity 2704 and secured into the housing 2706. The pods maycontain all layers needed for performing an immunoassay test, such asthat shown in FIG. 25, or they may be single layers, such as that shownin FIG. 24, where each layer is inserted down into the device 2700 tobuild a full immunoassay test.

Referring now to FIG. 28, there is illustrated a cross-sectional view ofone embodiment of an two-sided flow-through immunoassay testing device2800. The immunoassay testing device 2800 has a well 2802 having thereina plurality of orifices 2804. The plurality of orifices 2804 channelbiologic material deposited into the well 2804 onto differentimmunoassay pods 2806. The immunoassay pods 2806 may be pods such asthat described with respect to FIG. 25, or multiple pods such as thosein FIG. 24, stacked to create an immunoassay test. The device 2800 alsoincludes a results window 2808. Inside the device 2800, and between thewindow 2808 and the pods 2806, there are open sections 2810 below eachpod 2806 to allow a user to view the results of the tests as presentedon the reaction layers of the pods 2806 through the window 2808.

Referring now to FIG. 29A, there is illustrated a top view of anotherembodiment of a two-sided flow-through immunoassay device 2900. Thedevice 2900 includes a top housing section 2902, the center of which isa cross-shape that creates four open area or four quadrants. The fourquadrants server as a well 2904 for the device 2900.

Referring now to FIGS. 29B-D, there is illustrated cross-sectional viewsof the device 2900. There is again shown the top housing section 2902having a center prong 2904. Below the top housing section 2902 is a setof immunoassay test layers 2906 having a dividing layer 2908 on top. Thedividing layer 2908 may be made of cotton or paper or some other similarmaterial. The set of immunoassay test layers 2906 also has an indention2910 therein. When the top housing section 2902 is connected to thedevice after an immunoassay test is installed, the prong 2904 pushesdown on the dividing layer 2908, pushing the dividing layer 2908 downinto the indention 2910. This causes multiple sections of theimmunoassay test to be formed that are associated with the fourquadrants of the top section 2902, allowing for multiple tests to berun, with each test being under each quadrant, and preventing biologicmaterial from moving between quadrants due to the prong 2904 dividingthe quadrants from each other. A window 2912 in a bottom housing section2914 allows a user to view the results of the tests.

Referring to FIG. 29E, there is illustrated a top perspective view ofthe device 2900. There is shown the top housing section 2902 separatedfrom the bottom housing section 2914, allowing for the immunoassay testlayers to be placed within the device 2900.

The customizability of the devices shown in FIGS. 23A-29E allows forboth manufacturers and users of the devices to build custom immunoassaytesting devices. Manufacturers often have to build each immunoassay teststrip, laminate them, and insert them into a housing to be sold as thedevice. However, the devices disclosed herein allow for uniformcassettes to be on hand that can have pods inserted to custom build thetests, and allowing for layers to be stacked, without having topainstakingly build each test strip. Users also benefit since pods couldbe sold as separate replacements, allowing a user to buy a singlecassette, and periodically buy disposable pods to be used and thendiscarded after a test is complete, keeping the cassette for the nexttime the user needs to perform a test.

It should be understood that the drawings and detailed descriptionherein are to be regarded in an illustrative rather than a restrictivemanner, and are not intended to be limiting to the particular forms andexamples disclosed. On the contrary, included are any furthermodifications, changes, rearrangements, substitutions, alternatives,design choices, and embodiments apparent to those of ordinary skill inthe art, without departing from the spirit and scope hereof, as definedby the following claims. Thus, it is intended that the following claimsbe interpreted to embrace all such further modifications, changes,rearrangements, substitutions, alternatives, design choices, andembodiments.

What is claimed is:
 1. A two-sided flow through immunoassay device, thedevice comprising: a body including a top surface and a bottom surface;one or more sample wells disposed on the top surface of the body; one ormore orifices disposed vertically within the body, each of the one ormore orifices including an opening to one of the one or more samplewells, and the one or more orifices configured to receive a biologicsample; and one or more enclosed areas, each of the one or more enclosedareas disposed below one of the one or more orifices and configured toreceive the biologic sample from the one or more orifices, wherein eachof the one or more enclosed areas includes a set of immunoassay layers.2. The device of claim 1, further comprising a vacuum port disposed onthe bottom surface of the body and configured to receive a suctiondevice.
 3. The device of claim 2, wherein the suction device isconfigured to pull the biologic sample through the device.
 4. The deviceof claim 1, further comprising an alignment indicator disposed on thetop surface of the body.
 5. The device of claim 4, wherein the alignmentindicator triggers a mobile device to capture an image of the device. 6.The device of claim 1, wherein the set of immunoassay layers includes awicking layer disposed at a bottom of the set of immunoassay layers. 7.The device of claim 1, wherein the set of immunoassay layers includes aliquid permeable membrane layer, wherein the liquid permeable membranelayer allows liquid to enter further into the one or more orifices. 8.The device of claim 1, further comprising a test results window disposedon the bottom surface of the body.
 9. The device of claim 8, wherein theset of immunoassay layers includes an opaque layer that allows thebiologic sample to pass therethrough and prevents a user from seeingthrough the one or more sample wells to the test results window.
 10. Thedevice of claim 1, wherein the set of immunoassay layers includes areaction membrane, wherein the reaction membrane includes conjugates forbinding as a result of a reaction with the biologic sample.
 11. Thedevice of claim 10, wherein the reaction membrane includes test resultindicators that provide a qualitative test result of the reaction withthe biologic sample.
 12. The device of claim 1, wherein the set ofimmunoassay layers includes a blocking layer to filter the biologicsample.
 13. The device of claim 1, further comprising a first batterycompartment and a second battery compartment that provide a positive ornegative current to the set of immunoassay layers.
 14. The device ofclaim 1, wherein the set of immunoassay layers is disposed within ahousing.
 15. The device of claim 14, wherein the housing includes afirst lip disposed on a top of the set of immunoassay layers and asecond lip disposed on a bottom of the set of immunoassay layers. 16.The device of claim 15, wherein the first lip of the housing extendspartially over the top of the set of immunoassay layers such that acenter portion of the top of the set of immunoassay layers is uncoveredto allow the biologic sample to contact the set of immunoassay layers.17. The device of claim 16, wherein the second lip of the housingextends partially over the bottom of the set of immunoassay layers suchthat a center portion of the bottom of the set of immunoassay layers isuncovered to allow test results provided by the set of immunoassaylayers to be viewed through a test results window.
 18. The device ofclaim 17, wherein the housing has a front end and a back end, whereinthe front end and the back end are enclosed to hold the set ofimmunoassay layers together.
 19. The device of claim 18, wherein thehousing is configured to be removed from the one or more enclosed areasof the device.
 20. The device of claim 1, wherein the each of the one ormore enclosed areas prevent the biologic sample from entering anotherone of the one or more enclosed areas.